EP0012957B1 - Diagnostic means for the detection of proteolytic enzymes and indoxyl or thioindoxyl ester chromogens useful therefor, process for their preparation and their use in the preparation of diagnostic means - Google Patents

Description

The detection of leukocytes in body fluids, especially in the urine, plays an outstanding role in the diagnosis of diseases of the kidney and urogenital tract.

So far, this detection has been carried out by laboriously counting the leukocytes in the non-centrifuged urine or in the urine sediment.

It is common to both methods that only intact leukocytes are recorded. On the other hand, it is known that the speed of leukocyte lysis is subject to enormous fluctuations depending on the urinary environment; for example In strongly alkaline urines, a leukocyte half-life of only 60 minutes can be expected. The result is low leukocyte counts or, in the case of long urine retention times, false negative results.

Apart from the lysis error, the quantitative microscopic determination of the leukocytes in the non-centrifuged, homogenized urine in the counting chamber provides quite reliable values. In practice, however, this method is rarely used because it is tedious, tiring and time-consuming and requires the use of trained personnel.

The vast majority of leukocyte determinations in urine are carried out in medical practice using the so-called visual field method in urine sediment. To do this, the test material (sediment) must first be obtained by centrifugation. However, other components of the urine are also enriched, such as Salts and epithelial cells - can make microscopic counting of leukocytes considerably more difficult. Fluctuating sediment content, inhomogeneity of the sediment, as well as possibly different microscopic magnifications or different optical equipment of the microscopes mean that the usual information about the number of leukocytes per microscopic field of view can be affected by errors of several hundred percent.

The object of the present invention was therefore to provide a diagnostic agent with which the leukocytes in body fluids can be detected in a simple and easy-to-use manner and as quickly and completely as possible.

An enzymatic reaction lends itself to the principle of detection for such a leukocyte test, since the leukocytes have a broad spectrum of enzymes.

US Pat. No. 3,087,794 describes and claims a leukocyte detection which is carried out via the peroxidatic activity present in the granulocytic leukocytes. An absorbent carrier impregnated with hydrogen peroxide and an organic indicator, for example o-tolidine, indicates the presence of leukocytes by the formation of a colored oxidation product. However, such a test has decisive disadvantages: On the one hand, peroxidatic reactions using o-tolidine generally have a reducing effect on reducing substances in the urine, e.g. against ascorbic acid, a considerable susceptibility to interference. In addition, several references (see e.g. L. Mettler, Med. Welt 23, 399 [1972]) provide information on the instability of leukocyte peroxidase in the urinary environment, which gives rise to false negative results. The expected poor selectivity of this test towards erythrocytes is even more serious.

For several years, detection methods based on the esterolytic activity of the enzymes present in the systems to be determined have had a permanent place in histochemical and cytochemical enzymology (see e.g. A.G.E. Pearse, Histochemistry, Theoretical and Applied). In principle, colorless or weakly colored esters are used, which mostly break down into a colorless acid and also a colorless alcohol (phenol) component due to the enzymatic cleavage. The latter is then converted to colored products in a reaction following enzymatic saponification (e.g. coupling with diazonium salts or oxidative reactions).

For example, F. Schmalzl and H. Braunsteiner in Klin. Wschr. 46, 642 (1968) a specific cytochemical detection of leukocyte esterase with naphthol-AS-D-chloroacetate as substrate and a diazonium salt to form the colored azo compound.

Two-component systems of this type have not proven to be suitable for a diagnostic means for the quick and simple detection of leukocytes in body fluids, such as in urine, since it is known that many compounds found in urine, such as urobilinogen, stercobilinogen, bilirubin and others, with diazonium salts react. In addition, this proof is far too insensitive. For example, samples of 5000 leukocytes / _I ll no reaction.

British patent 11 28 371 describes and claims a diagnostic agent for the detection of hydrolytic enzymes in body fluids. Here, an absorbent carrier is impregnated with colorless indoxyl or thioindoxyl esters and optionally a buffer and an oxidizing agent. In the presence of hydrolytic enzymes, free indoxyl or thioindoxyl is formed from the esters and from this deeply colored indigo or thioindigo by the action of atmospheric oxygen or oxidizing agent. The compounds claimed in this patent are not useful for a leukocyte test because they show no reaction even at 10,000 leukocytes / µI.

To date, there is no test strip on the market that it allows easy and quick detection of leukocytes, although the detection of leukocytes in the urine is one of the most frequently performed clinical tests.

Surprisingly, it has now been found that stable and rapidly indicating diagnostic agents with which leukocytes can be readily detected in body fluids are obtained when substrates for the detection of the esterases (proteases), indoxyl or thioindoxyl amino acid esters or present in the neutrophilic leukocyte granulocytes -Peptide esters can be used. In addition, it was shown that these substrates are also excellent for the general detection of proteolytic enzymes, e.g. of elastase, chymotrypsin or trypsin in purely aqueous solutions or also in body fluids, e.g. Plasma, serum, cerebrospinal fluid, pancreatic secretions or aqueous stool extracts are suitable.

in der

• R_i, R₂, R₃, R₄, die gleich oder verschieden sein können, jeweils Wasserstoff oder Halogen, eine niedere Alkyl-, niedere Alkoxy-, Aryl-, Aralkyl-, Aralkoxy-, eine Hydroxy-, eine Carboxy-, eine Carboxy-nieder-Alkoxy-, eine Aralkoxycarbonyl-, eine Aralkoxycarbonyl-nieder-Alkoxy-, eine Nitro-oder eine niedere Acylaminogruppe oder jeweils zwei benachbarte Substituenten einen gegebenenfalls durch Halogen substituierten benzoannellierten Rest,
• X ein Schwefelatom oder eine gegebenenfalls durch einen niederen Alkyl-, einen Aryl-, einen Aralkyl-oder einen Acylrest substituierte Iminogruppe,
• A einen Aminosäure- oder einen Peptidrest, B eine in der Peptidchemie übliche oder davon abgeleitete Stickstoffschutzgruppe bedeutet, eingesetzt werden.

The present invention therefore relates to a diagnostic agent for the detection of proteolytic enzymes, in particular for the detection of the proteases present in the leukocytes in body fluids, consisting of an absorbent carrier, a film layer, a powder mixture, a lyophilisate, a solution or a reagent tablet, containing one or several chromogens and conventional additives, characterized in that as chromogens indoxyl and / or thioindoxyl amino acid esters and / or peptide esters of the general formula I

in the

• R _i , R ₂ , R ₃ , R ₄ , which can be the same or different, each hydrogen or halogen, a lower alkyl, lower alkoxy, aryl, aralkyl, aralkoxy, a hydroxy, a carboxy, a carboxy-lower alkoxy, an aralkoxycarbonyl, an aralkoxycarbonyl-lower alkoxy, a nitro or a lower acylamino group or two adjacent substituents each an optionally halogen-substituted benzo-fused radical,
• X is a sulfur atom or an imino group which is optionally substituted by a lower alkyl, an aryl, an aralkyl or an acyl radical,
• A is an amino acid or a peptide residue, B is a nitrogen protecting group which is customary in peptide chemistry or is derived therefrom.

Another object of the present invention is the use of indoxyl and / or thioindoxyl amino acid esters and / or peptide esters of the general formula for the preparation of diagnostic agents for the detection of proteolytic enzymes, in particular the proteases present in the leukocytes in body fluids.

All indoxyl and thioindoxyl amino acid esters and peptide esters of the general formula are new compounds.

The present invention therefore also relates to the indoxyl and thioindoxyl amino acid esters and peptide esters of the general formula I and processes for their preparation.

The new indoxyl and thioindoxyl amino acid esters and peptide esters of the general formula I can be prepared by methods known per se from peptide chemistry.

in der R_i, R₂, R₃, R₄ und X die oben angegebene Bedeutung haben, mit Aminosäuren und Peptiden der allgemeinen Formel III

in der A und B die oben angegebene Bedeutung haben, beziehungsweise mit geeigneten reaktiven Derivaten davon, umgesetzt.Preferably, the corresponding indoxyl or thioindoxyl compounds of the general formula II are known

in which R _i , R ₂ , R ₃ , R ₄ and X have the meaning given above, with amino acids and peptides of the general formula III

in which A and B have the meaning given above, or with suitable reactive derivatives thereof.

As reactive derivatives e.g. the acid chlorides, or the mixed anhydrides usually used in peptide synthesis, for example with ethyl chloroformate or active ester.

The indoxyl and thioindoxyl compounds of the general formula II, as well as the amino acids and peptides of the general formula III are known substances (see, for example, P. Friedländer, Advances in Tar Dye Manufacturing and Related Industries, Vol. 3-20 and Houben-Weyl, Methods of organic chemistry, Vol. 15/1), or can be prepared in analogy to known compounds.

Halogen in the definition of R _1, R ₂ , R ₃ and R ₄ is to be understood as fluorine, chlorine, bromine and iodine, preferably chlorine and bromine.

The “lower alkoxy group” in the definition of R ₁ , R ₂ , R ₃ and R ₄ , and the “lower alkyl group” of R ₁ , R ₂ , R ₃ , R ₄ and X contain 1 to 5, preferably 1 to 3 Carbon atoms, with the methoxy and the methyl group being particularly preferred.

An “aralkoxy group” in the definition of R _i , R ₂ , R ₃ and R ₄ , and an “aralkyl group” in the definition of R ₁ , R ₂ , R ₃ , R ₄ and X are, for example, by oxy-lower To understand alkyl or lower alkyl radicals substituted phenyl and naphthyl groups, the alkyl radical containing 1 to 5, preferably 1 to 3, carbon atoms. The benzyloxy and the benzyl radical are particularly preferred.

The “lower acylamino group” of R _i , R ₂ , R ₃ and R ₄ are the amide groups of the lower aliphatic carboxylic acids having 1 to 5, preferably 1 to 3, carbon atoms. The acetylamino radical is particularly preferred. The “acyl radical” in the definition of X is the radicals of aliphatic carboxylic acids with 1 to 5, preferably 1 to 3, carbon atoms, or also aromatic carboxylic acids, such as, for example, benzoic or naphthoic acids. Acetyl and benzoyl radicals are particularly preferred.

An “aryl radical” in the definition of R ₁ , R ₂ , R ₃ , R ₄ and X is preferably to be understood as the phenyl or naphthyl group.

The “amino acid residue” in the definition of A is preferably the residues of the natural a-amino acids in their L or D form or also in their racemic form. The residues of glycine, alanine, valine, leucine, isoleucine, phenylalanine and tyrosine are particularly preferred. Any free hydroxyl groups present can be acylated, preferably acetylated.

Under a "peptide residue" in the definition of A are e.g. To understand di-, tri-, tetra- and pentapeptides, preferably di- and tripeptides, the amino acid components mentioned preferably being used as the amino acid components.

As "nitrogen protecting group common in peptide chemistry" in the definition of B are e.g. To understand acyl, oxycarbonyl, thiocarbonyl, sulfonyl, sulfenyl, vinyl, cyclohexenyl, phosphoryl or carbamoyl groups.

The indoxyl or thioindoxyl amino acid esters and peptide esters of the general formula used according to the invention as chromogens are used in concentrations of 10 to ⁴ to 1 mol / liter, preferably 10 to ³ to 10 mol / liter of impregnating solution, coating composition or liquid to be examined.

Another component of the diagnostic agent for the detection of proteolytic enzymes and in particular the leukocyte proteases is a suitable buffer system. In addition come e.g. Phosphate, borate, barbiturate, tris (hydroxymethyl) aminomethane - (= tris -) -, 2-amino-2-methyl-propanediol-1,3 - (= amediol) - or amino acid buffer in question, the pH and capacity must be selected so that a pH of 6-10, preferably 7-9, is established in the measurement solution or on the test strip.

In addition, in the production of the diagnostic agent according to the invention for the detection of proteolytic enzymes, in particular the leukocyte proteases in body fluids, additional oxidizing agents can be used in order to convert the indoxyl or thioindoxyl compounds, which primarily result from the enzymatic reaction, to the colored indigo or thioindigo compounds. Implement substances. These oxidizing agents, such as, for example, potassium hexacyanoferrate III, potassium bromate, potassium chromate, phenazine methosulfate or tetrazolium salts, are used in concentrations of 10 to ⁴ to 1 mol / liter, preferably 10 to 10 mol / liter, of impregnating solution, coating composition or liquid to be examined used.

A further component of a diagnostic agent for the detection of proteolytic enzymes, in particular leukocyte proteases, can be a wetting agent, since this results in somewhat shorter reaction times and sometimes. brilliant colors can be achieved. Nonionic but also amphoteric, cationic or anionic wetting agents are preferably used in concentrations of 0.05-2%, preferably 0.1-1%.

For the preparation of the agent according to the invention e.g. absorbent carrier, preferably filter paper, cellulose or synthetic fiber fleece, with solutions of the necessary reagents (substrate, buffer, optionally wetting agent, oxidizing agent, etc.) usually used for the production of test strips in volatile solvents, such as e.g. Water, methanol, ethanol or acetone, impregnated. This is conveniently done in two separate steps: First, impregnation with an aqueous solution that contains the buffer and other water-soluble additives. It is then impregnated with a solution of the protease substrates of the general formula I. In special cases the reverse impregnation sequence can also be used.

The finished test papers can be used as such or glued to handles in a manner known per se or preferably sealed between plastics and fine-meshed networks in accordance with DBP 21 18 455.

To produce film-coated test strips, all reagents are dissolved in the solution or dispersion of a film-forming substance, e.g. Polyvinyl ester or polyamide, registered and mixed homogeneously. The mixture is spread in a thin layer on a plastic carrier and dried. The film-coated test strips according to the invention are cut after drying and can be used as such or glued to handles in a manner known per se or e.g. between plastics and fine-mesh networks according to DBP 21 18 455.

The diagnostic agent according to the invention for the detection of proteolytic enzymes, in particular the leukocyte proteases, in the form of powder mixtures or reagent tablets can be produced by adding the usual components of the test to the components of the test and granulating them. Additives of this type are e.g. Carbohydrates, e.g. Mono-, oligo- or polysaccharides, or sugar alcohols, e.g. Mannitol, sorbitol or xylitol, or other soluble inert compounds such as polyethylene glycols or polyvinylpyrrolidone. The powder mixtures or reagent tablets generally have a final weight of approximately 50-200 mg, preferably 50-80 mg.

For the production of lyophilisates with a total weight of about 5-20 mg, preferably about 10 mg, a solution is freeze-dried which, in addition to all the Re Agents customary scaffolders, such as polyvinyl pyrrolidone, and possibly other fillers, such as mannitol, sorbitol or xylitol, contains.

The diagnostic agent according to the invention in the form of a solution preferably contains all the reagents required for the test. Water or mixtures of water with a water-soluble organic solvent, such as e.g. Methanol, ethanol, acetone or dimethylformamide, in question. For reasons of durability, it can be advantageous to distribute the reagents required for the test over two or more solutions, which are only combined during the actual examination.

The diagnostic agents thus prepared make it possible to quickly and easily detect the presence of proteolytic enzymes, in particular the leukocyte proteases, after immersion in the body fluid to be examined or after addition to the relevant body fluid, by color formation that can be visual or photometric, e.g. remission photometric or in the cuvette. Since the activity of the leukocyte proteases per cell can be regarded as an essentially constant variable, the leukocyte concentration of the examined body fluid can be determined from the intensity of the color formation. Both intact and lysed leukocytes are detected with the diagnostic agent according to the invention, since the activity of the leukocyte proteases is fully retained even after the leukocytes have been lysed. A lysis error therefore does not occur.

example 1

Filter paper (eg Schleicher + Schüll 23 SL) is successively impregnated with the following solutions and then dried at 60 ° C.

• 5000 Leukozyten/_Ill Harn in ca. 2 Minuten
• 1000 Leukozyten/µl Harn in ca. 6 Minuten
• 500 Leukozyten/µl Harn in ca. 10 Minuten
• 200 Leukozyten/µl Harn in ca. 15 Minuten.

A colorless test paper is obtained which, when immersed in urine containing leukocytes, changes color from light turquoise to blue depending on the leukocyte concentration. The following can be demonstrated:

• 5000 leukocytes / _I ll urine in about 2 minutes
• 1000 leukocytes / µl urine in about 6 minutes
• 500 leukocytes / µl urine in about 10 minutes
• 200 leukocytes / µl urine in approx. 15 minutes.

The sensitivity of the test is around 200 leukocytes / µl. The assessment can also be carried out using reflectance photometry at 620 nm.

• 1.1. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-4-methyl-indol
• 1.2. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-5-methyl-indol
• 1.3. 3-[N-(Benzyloxycarbonyl) -L-alanyloxy]-6-methyl-indol
• 1.4. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-7-methyl-indol
• 1.5. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-4,7-dimethyl-indol
• 1.6. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-4-chlor-indol
• 1.7. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-5-brom-indol
• 1.8. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-6-chlor-indol Verfärbung: farblos nach purpur
• 1.9. 3-[N-(Toluol- 4'-sulfonyl)-L-alanyloxy]-4-chlor-5-brom-indol
• 1.10. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4,5,7-trichlor-indol
• 1.11. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4-chlor-5-brom-7-methyl-indol
• 1.12. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-hydroxy-indol
• 1.13. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-5-methoxy-indol
• 1.14. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-benzyloxy-indol .
• 1.15. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4-carboxy-indol
• 1.16. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4-benzyloxycarbonyl-indol
• 1.17. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-(carboxy-methoxy)-indol
• 1.18. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxyl-5-(benzyloxycarbonyl-methoxy)-indol
• 1.19. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxyl-6-nitro-indol
• 1.20. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-6-acetylamino-indol
• 1.21. 3-[N-(Toluot-4'-sulfonyl)-L-alanyloxy]-benzo[g]-indol Verfärbung: farblos nach grün.
• 1.22. 1-Methyl-3-[N-(benzyloxycarbonyl)-L-alanyloxy]-indol Verfärbung: farblos nach grün.
• 1.23 1-Benzyl-3-[N-(toluol-4'-sulfonyl)- L-alanyloxy]-indol Verfärbung: farblos nach grün.
• 1.24. 1-Phenyl-3-[N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol Verfärbung: farblos nach blaugrün.
• 1.25 1-Acetyl-3-[N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol Verfärbung: farblos nach rot.
• 1.26. 1-Benzoyl-3-(N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol Verfärbung: farblos nach violett.
• 1.27. 3-[N-(Benzyloxycarbonyl)-glycyloxy]-indol
• 1.28. 3-[N-(Toluol-4'-sulfonyl)-glycyloxy]-indol
• 1.29. 3-[N-(Toluol-2'-sulfonyl)-L-alanyloxy]-indol
• 1.30. 3-[N-(Toluol-3'-sulfonyl)-L-alanyloxy]-indol
• 1.31. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-indol
• 1.32. 3-[N-(Toluoi-4'-sulfonyl)-D-alanyloxy]-indol
• 1.33. 3-[N-(Benzyloxycarbonyl)-D,L-alanyloxy]-indol
• 1.34. 3-[N-Benzyloxycarbonyl)-L-valyloxy]-indol
• 1.35. 3-[N-(Toluol-4'-sulfonyl)-L-valyloxy]-indol
• 1.36. 3-[N-(Benzyloxycarbonyl)-L-leucyloxy]-indol
• 1.37. 3-[N-(Benzyloxycarbonyl)-L-isoleucyloxy]-indol
• 1.38. 3-[N-(Benzyloxycarbonyl)-L-phenylalanyloxy]-indol
• 1.39. 3-(N-(Toluol-4'-sulfonyl)-L-phenylalanyloxy]-indol
• 1.40. 3-[N-Acetyl-L-tyrosyloxy]-indol
• 1.41. 3-[N-Benzoyl-L-tyrosyloxy]-indol
• 1.42. 3-[N-(Benzyloxycarbonyl)-L-tyrosyloxy]-indol
• 1.43. 3-[N-(Toluol-4'-sulfonyl)-L-tyrosyloxy]-indol
• 1.44. 3-[N-(Toluol-4'-sulfonyl)-0-acetyl-L-tyrosyloxy]-indol
• 1.45. 3-[N-(Benzyloxycarbonyl)-L-alanyl-L-alanyloxy]-indol
• 1.46. 3-[N-(Toluol-4'-sulfonyl)-D-alanyl-L-alanyloxy]-indol
• 1.47. 3-[N-(Benzyloxycarbonyl)-L-alanyl-L-alanyl-L-alanyloxy]-indol
• 1.48. 3-[N-(Toluol-4'-sulfonyl)-D-alanyl-D-alanyl-L-alanyloxy]-indol
• 1.49. 3-[N-Formyl-L-alanyloxy]-indol
• 1.50. 3-[N-Acetyl-L-alanyloxy]-indol
• 1.51. 3-[N-Succinyl-L-alanyloxy]-indol
• 1.52. 3-[N-Benzoyl-D,L-alanyloxy]-indol
• 1.53. 3-[N-Phthaloyl-L-alanyloxy]-indol
• 1.54. 3-[N-(Ethoxycarbonyl)-L-alanyloxy]-indol
• 1.55. 3-[N-(tert.-Butyloxycarbonyl)-L-alanyloxy]-indol
• 1.56. 3-[N-(3',6'-Dioxa-n-heptyloxycarbonyl)-L-alanyloxy]-indol
• 1.57. 3-[N-(Cyclohexyloxycarbonyl)-L-alanyloxy]-indol
• 1.58. 3-[N-(Phenyloxycarbonyl)-L-alanyloxy]-indol
• 1.59. 3-[N-(4'-Methyl-benzyloxycarbonyl)-L-alanyloxy]-indol
• 1.60. 3-[N-(4'-Methoxy-benzyloxycarbonyl)-L-alanyloxy]-indol
• 1.61. 3-[N-(4'-Nitro-benzyloxycarbonyl)-L-alanyloxy]-indol
• 1.62. 3-[N-(N'-(Piperidino)-oxycarbonyl)-L-alanyloxy]-indol
• 1.63. 3-[N-(Furyl-(2')-methoxycarbonyl)-L-alanyloxy]-indol
• 1.64. 3-[N-(Thienyl-(2')-methoxycarbonyl)-L-alanyloxy]-indol
• 1.65. 3-[N-(Benzylthiocarbonyl)-L-alanyloxy]-indol
• 1.66. 3-[N-(Methansulfonyl)-L-alanyloxy]-indol
• 1.67. 3-[N-(Benzylsulfonyl)-L-alanyloxy]-indol
• 1.68. 3-[N-(Benzolsulfonyl)-L-alanyloxy]-indol
• 1.69. 3-[N-(4'-Brom-benzolsulfonyl)-L-alanyloxy]-indol
• 1.70. 3-[N-(4'-Nitro-benzolsulfonyl)-L-alanyloxy]-indol
• 1.71. 3-[N-(4'-Dimethylamino-benzolsulfonyl)-L-alanyloxy]-indol
• 1.72. 3-[N-(4'-Acetylamino-benzolsulfonyl)-L-alanyloxy]-indol
• 1.73. 3-[N-(4'-n-Butyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.74. 3-[N-(4'-tert.-Butyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.75. 3-[N-(4'-n-Octyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.76. 3-[N-(4'-Hydroxy-benzolsulfonyl)-L-alanyloxy]-indol
• 1.77. 3-[N-(4'-Methoxy-benzolsulfonyl)-L-alanyloxy]-indol
• 1.78. 3-[N-(4'-Benzyloxy-benzolsulfonyl)-L-alanyloxy]-indol
• 1.79. 3-[N-(4'-(2"-Hydroxy-ethoxy)-benzolsulfonyl)-L-alanyloxy]-indol
• 1.80. 3-[N-(4'-(3"-Oxa-5"-hydroxy-n-pentyl- oxy)-benzolsulfonyl)-L-alanyloxy]-indol
• 1.81. 3-[N-(4'-(3",6"-Di-oxa-n-heptyloxy)-benzolsulfonyl)-L-alanyloxy]-indol
• 1.82. 3-[N-( 4'-(2"-Hydroxy-ethyl)-benzol- sulfonyl)-L-alanyloxy]-indol
• 1.83. 3-[N-(4'-(2"-{4"'-Nitro-benzyloxy}- ethyl)-benzolsulfonyl)-L-alanyloxy]-indol
• 1.84. 3-[N-(4'-(2"-Chlor-ethyl)-benzolsulfo- nyl)-L-alanyloxy]-indol
• 1.85. 3-[N-(4'-Acetyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.86. 3-[N-(4'-Cyano-benzolsulfonyl)-L-. alanyloxy]-indol
• 1.87. 3-[N-(4'-Carboxy-benzolsulfonyl)-L-alanyloxy]-indol
• 1.88. 3-[N-(4'-Methoxycarbonyl-benzolsulfo- nyl)-L-alanyloxy]-indol
• 1.89. 3-[N-(4'-Benzyloxycarbonyl-benzolsulfo- nyl)-L-alanyloxy]-indol
• 1.90. 3-[N-(4'-Carbamoyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.91. 3-[N-(4'-(Dimethyl-carbamoyl)-benzolsulfonyl)-L-alanyloxy]-indol
• 1.92. 3-[N-(4'-Carboxymethyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.93. 3-[N-(4'-Carboxymethoxy-benzolsulfonyl)-L-alanyloxy]-indol
• 1.94. 3-[N-(4'-Carboxymethylamino-benzolsul- fonyl)-L-alanyloxy]-indol
• 1.95. 3-[N-(4'-(Benzyloxycarbonyl-methyl- amino)-benzolsulfonyl)-L-alanyloxy]-indol
• 1.96. 3-[N-(4'-FIuor-benzolsufonyl)-L-alanyloxy]-indol
• 1.97. 3-[N-(4'-Fluorsulfonyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.98. 3-[N-(4'-Sulfamoyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.99. 3-[N-Methyl-N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol
• 1.100. 3-[N-Acetyl-N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol
• 1.101. 3-[N-(2',4',6'-Trimethyl-benzolsulfonyl)-L-alanyloxy]-indol
• 1.102. 3-[N-(Biphenyl-4'-sulfonyl)-L-alanyloxy]-indol
• 1.103. 3-[N-(Naphthalin-2'-sulfonyl)-L-alanyloxyl-indol
• 1.104. 3-[N-(4'-Acetylamino-naphthalin-1'-sulfonyl)-L-alanyloxy]-indol
• 1.105. 3-[N-(5'-Dimethylamino-naphthalin- l'-sulfonyl)-L-alanyloxyl-Indol
• 1.106. 3-[N-(Chinolin-8'-sulfonyl)-L-alanyloxy]-indol
• 1.107. 3-[N-(F^>yridin-3'-sulfonyl)-L-alanyloxy]-indol
• 1.108. 3-[N-(2'-Nitro-benzolsulfonyl)-L-alanyloxy]-indol Verfärbung: gelb nach grün.
• 1.109. 3-[N-(1'-Methyl-2'-benzoyl-vinyl)-L-alanyloxy]-indol
• 1.110. 3-[N-(5',5'-Dimethyl-3'-oxo-cyclohexen-1'-yl)-L-alanyloxy]-indol
• 1.111. 3-[N(Diphenylcarbamoyl)-L-alanyloxy]-indol
• 1.112. 3-[N-(Di-(4'-nitrobenzyl)-phosphoryl)-L-alanyloxy]-indol
• 1.113. 3-[N-(Di-(4'-brombenzyl)-phosphoryl)-L-alanyloxy]-indol
• 1.114. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-benzyl-i ndol
• 1.115. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-phenyl-indol
• 1.116. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-methoxy-indol

Test papers with similar properties (sensitivities: 200-2,000 leukocytes / µl are obtained if, instead of 3- [N- (benzyloxycarbonyl) -L-alanyloxy] indole, the following substrates are used, although - if not specifically mentioned - also Have light turquoise to blue discoloration of the otherwise colorless test papers observed when immersed in leukocyte-containing urine:

• 1.1. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -4-methyl-indole
• 1.2. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -5-methyl-indole
• 1.3. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -6-methyl-indole
• 1.4. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -7-methyl-indole
• 1.5. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -4,7-dimethyl-indole
• 1.6. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -4-chloroindole
• 1.7. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -5-bromo-indole
• 1.8. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -6-chloro-indole Discoloration: colorless to purple
• 1.9. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-chloro-5-bromo-indole
• 1.10. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4,5,7-trichloroindole
• 1.11. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-chloro-5-bromo-7-methyl-indole
• 1.12. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-hydroxy-indole
• 1.13. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -5-methoxy-indole
• 1.14. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-benzyloxy-indole.
• 1.15. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-carboxy-indole
• 1.16. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-benzyloxycarbonyl-indole
• 1.17. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5- (carboxymethoxy) indole
• 1.18. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxyl-5- (benzyloxycarbonyl-methoxy) indole
• 1.19. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxyl-6-nitro-indole
• 1.20. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -6-acetylamino-indole
• 1.21. 3- [N- (Toluot-4'-sulfonyl) -L-alanyloxy] -benzo [g] -indole Discoloration: colorless to green.
• 1.22. 1-Methyl-3- [N- (benzyloxycarbonyl) -L-alanyloxy] indole Discoloration: colorless to green.
• 1.23 1-Benzyl-3- [N- (toluene-4'-sulfonyl) - L-alanyloxy] indole Discoloration: colorless to green.
• 1.24. 1-phenyl-3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] indole Discoloration: colorless to blue-green.
• 1.25 1-Acetyl-3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] indole Discoloration: colorless to red.
• 1.26. 1-Benzoyl-3- (N- (toluene-4'-sulfonyl) -L-alanyloxy] indole Discoloration: colorless to violet.
• 1.27. 3- [N- (benzyloxycarbonyl) glycyloxy] indole
• 1.28. 3- [N- (toluene-4'-sulfonyl) glycyloxy] indole
• 1.29. 3- [N- (toluene-2'-sulfonyl) -L-alanyloxy] indole
• 1.30. 3- [N- (toluene-3'-sulfonyl) -L-alanyloxy] indole
• 1.31. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] indole
• 1.32. 3- [N- (Toluoi-4'-sulfonyl) -D-alanyloxy] indole
• 1.33. 3- [N- (Benzyloxycarbonyl) -D, L-alanyloxy] indole
• 1.34. 3- [N-benzyloxycarbonyl) -L-valyloxy] indole
• 1.35. 3- [N- (toluene-4'-sulfonyl) -L-valyloxy] indole
• 1.36. 3- [N- (Benzyloxycarbonyl) -L-leucyloxy] indole
• 1.37. 3- [N- (Benzyloxycarbonyl) -L-isoleucyloxy] indole
• 1.38. 3- [N- (Benzyloxycarbonyl) -L-phenylalanyloxy] indole
• 1.39. 3- (N- (toluene-4'-sulfonyl) -L-phenylalanyloxy] indole
• 1.40. 3- [N-acetyl-L-tyrosyloxy] indole
• 1.41. 3- [N-benzoyl-L-tyrosyloxy] indole
• 1.42. 3- [N- (Benzyloxycarbonyl) -L-tyrosyloxy] indole
• 1.43. 3- [N- (toluene-4'-sulfonyl) -L-tyrosyloxy] indole
• 1.44. 3- [N- (toluene-4'-sulfonyl) -0-acetyl-L-tyrosyloxy] indole
• 1.45. 3- [N- (Benzyloxycarbonyl) -L-alanyl-L-alanyloxy] indole
• 1.46. 3- [N- (toluene-4'-sulfonyl) -D-alanyl-L-alanyloxy] indole
• 1.47. 3- [N- (Benzyloxycarbonyl) -L-alanyl-L-alanyl-L-alanyloxy] indole
• 1.48. 3- [N- (toluene-4'-sulfonyl) -D-alanyl-D-alanyl-L-alanyloxy] indole
• 1.49. 3- [N-formyl-L-alanyloxy] indole
• 1.50. 3- [N-acetyl-L-alanyloxy] indole
• 1.51. 3- [N-succinyl-L-alanyloxy] indole
• 1.52. 3- [N-benzoyl-D, L-alanyloxy] indole
• 1.53. 3- [N-phthaloyl-L-alanyloxy] indole
• 1.54. 3- [N- (Ethoxycarbonyl) -L-alanyloxy] indole
• 1.55. 3- [N- (tert-Butyloxycarbonyl) -L-alanyloxy] indole
• 1.56. 3- [N- (3 ', 6'-Dioxa-n-heptyloxycarbonyl) -L-alanyloxy] indole
• 1.57. 3- [N- (Cyclohexyloxycarbonyl) -L-alanyloxy] indole
• 1.58. 3- [N- (phenyloxycarbonyl) -L-alanyloxy] indole
• 1.59. 3- [N- (4'-Methyl-benzyloxycarbonyl) -L-alanyloxy] indole
• 1.60. 3- [N- (4'-Methoxy-benzyloxycarbonyl) -L-alanyloxy] indole
• 1.61. 3- [N- (4'-Nitro-benzyloxycarbonyl) -L-alanyloxy] indole
• 1.62. 3- [N- (N '- (Piperidino) oxycarbonyl) -L-alanyloxy] indole
• 1.63. 3- [N- (Furyl- (2 ') methoxycarbonyl) -L-alanyloxy] indole
• 1.64. 3- [N- (Thienyl- (2 ') methoxycarbonyl) -L-alanyloxy] indole
• 1.65. 3- [N- (Benzylthiocarbonyl) -L-alanyloxy] indole
• 1.66. 3- [N- (methanesulfonyl) -L-alanyloxy] indole
• 1.67. 3- [N- (benzylsulfonyl) -L-alanyloxy] indole
• 1.68. 3- [N- (benzenesulfonyl) -L-alanyloxy] indole
• 1.69. 3- [N- (4'-Bromo-benzenesulfonyl) -L-alanyloxy] indole
• 1.70. 3- [N- (4'-Nitro-benzenesulfonyl) -L-alanyloxy] indole
• 1.71. 3- [N- (4'-Dimethylamino-benzenesulfonyl) -L-alanyloxy] indole
• 1.72. 3- [N- (4'-Acetylamino-benzenesulfonyl) -L-alanyloxy] indole
• 1.73. 3- [N- (4'-n-Butyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.74. 3- [N- (4'-tert-Butyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.75. 3- [N- (4'-n-Octyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.76. 3- [N- (4'-Hydroxy-benzenesulfonyl) -L-alanyloxy] indole
• 1.77. 3- [N- (4'-Methoxy-benzenesulfonyl) -L-alanyloxy] indole
• 1.78. 3- [N- (4'-Benzyloxy-benzenesulfonyl) -L-alanyloxy] indole
• 1.79. 3- [N- (4 '- (2 "-hydroxy-ethoxy) -benzenesulfonyl) -L-alanyloxy] indole
• 1.80. 3- [N- (4 '- (3 "-Oxa-5" -hydroxy-n-pentyl-oxy) -benzenesulfonyl) -L-alanyloxy] indole
• 1.81. 3- [N- (4 '- (3 ", 6" -Di-oxa-n-heptyloxy) benzenesulfonyl) -L-alanyloxy] indole
• 1.82. 3- [N- (4 '- (2 "-hydroxy-ethyl) -benzenesulfonyl) -L-alanyloxy] indole
• 1.83. 3- [N- (4 '- (2 "- {4"' - Nitro-benzyloxy} ethyl) -benzenesulfonyl) -L-alanyloxy] indole
• 1.84. 3- [N- (4 '- (2 "Chloro-ethyl) benzenesulfonyl) -L-alanyloxy] indole
• 1.85. 3- [N- (4'-Acetyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.86. 3- [N- (4'-Cyano-benzenesulfonyl) -L-. alanyloxy] indole
• 1.87. 3- [N- (4'-Carboxy-benzenesulfonyl) -L-alanyloxy] indole
• 1.88. 3- [N- (4'-Methoxycarbonyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.89. 3- [N- (4'-Benzyloxycarbonyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.90. 3- [N- (4'-Carbamoyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.91. 3- [N- (4 '- (Dimethyl-carbamoyl) benzenesulfonyl) -L-alanyloxy] indole
• 1.92. 3- [N- (4'-Carboxymethyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.93. 3- [N- (4'-Carboxymethoxy-benzenesulfonyl) -L-alanyloxy] indole
• 1.94. 3- [N- (4'-Carboxymethylamino-benzenesulfonyl) -L-alanyloxy] indole
• 1.95. 3- [N- (4 '- (Benzyloxycarbonyl-methylamino) -benzenesulfonyl) -L-alanyloxy] indole
• 1.96. 3- [N- (4'-fluorobenzenesulfonyl) -L-alanyloxy] indole
• 1.97. 3- [N- (4'-Fluorosulfonyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.98. 3- [N- (4'-Sulfamoyl-benzenesulfonyl) -L-alanyloxy] indole
• 1.99. 3- [N-Methyl-N- (toluene-4'-sulfonyl) -L-alanyloxy] indole
• 1,100. 3- [N-acetyl-N- (toluene-4'-sulfonyl) -L-alanyloxy] indole
• 1,101. 3- [N- (2 ', 4', 6'-trimethyl-benzenesulfonyl) -L-alanyloxy] indole
• 1,102. 3- [N- (Biphenyl-4'-sulfonyl) -L-alanyloxy] indole
• 1,103. 3- [N- (naphthalene-2'-sulfonyl) -L-alanyloxyl indole
• 1,104. 3- [N- (4'-Acetylamino-naphthalene-1'-sulfonyl) -L-alanyloxy] indole
• 1,105. 3- [N- (5'-Dimethylamino-naphthalene-l'-sulfonyl) -L-alanyloxyl indole
• 1,106. 3- [N- (Quinoline-8'-sulfonyl) -L-alanyloxy] indole
• 1.107. 3- [N- (F ^> yridin-3'-sulfonyl) -L-alanyloxy] indole
• 1,108. 3- [N- (2'-Nitro-benzenesulfonyl) -L-alanyloxy] indole Discoloration: yellow to green.
• 1,109. 3- [N- (1'-Methyl-2'-benzoyl-vinyl) -L-alanyloxy] indole
• 1,110. 3- [N- (5 ', 5'-Dimethyl-3'-oxo-cyclohexen-1'-yl) -L-alanyloxy] indole
• 1,111. 3- [N (Diphenylcarbamoyl) -L-alanyloxy] indole
• 1,112. 3- [N- (Di- (4'-nitrobenzyl) phosphoryl) -L-alanyloxy] indole
• 1,113. 3- [N- (Di- (4'-bromobenzyl) phosphoryl) -L-alanyloxy] indole
• 1,114. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-benzyl-indole
• 1,115. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-phenyl-indole
• 1,116. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-methoxy-indole

Example 2

Filter paper (eg Schleicher + Schüll 23 SL) is successively impregnated with the following solutions and then dried at 60 ° C.

A yellow-colored test paper is obtained, which turns red when immersed in leukocyte-containing urine.

The sensitivity of the test is around 1000 leukocytes / µl urine.

The evaluation can also be carried out at 576 nm using reflectance photometry.

The substrates of Example 1 are treated with oxidizing agents, such as, for example, the potassium hexacyano ferrate III used above, or e.g. with potassium bromate, potassium chromate, phenazine methosulfate or tetrazolium salts, test papers are obtained which, in part, compared to analog test papers without oxidizing agents. have slightly reduced response times.

Example 3

Filter paper (eg Schleicher + Schüll 23 SL) is successively impregnated with the following solutions and then dried at 60 ° C.

A colorless test paper is obtained, which changes color from light turquoise to blue when immersed in leukocyte-containing urine, depending on the concentration of leukocytes. Compared to the recipe of Example 1, the reaction times are somewhat shorter and the colors are slightly more brilliant.

Also with the other substrates of Examples 1 and 2, wetting agents such as the nonylphenol polyglycol ether (nonionic) used above, but also e.g. with cocosimidazoline compounds (amphoteric) or benzyltrimethylammonium chloride (cationic) or sodium sulfonato-dodecylbenzene (anionic), obtained test papers, some of which are comparable to analog test papers without wetting agents. have slightly reduced response times and somewhat more brilliant colors.

Example 4

• 1 ml Lösung 1
• 1 ml Lösung 2
• 2 ml leukozytenhaltiger Harn

The following are mixed in a test tube:

• 1 ml solution 1
• 1 ml solution 2
• 2 ml of urine containing leukocytes

The mixture gradually changes color - depending on the leukocyte concentration - from light green to deep blue.

After standing for about 10 minutes at room temperature, the leukocyte concentration is determined visually using reference colors or photometrically, e.g. in 1 cm cuvettes at 620 nm.

The sensitivity of the test is around 200 leukocytes / µl urine.

Test tube or cuvette tests with similar sensitivities (200-1000 leukocytes / µl urine) can also be carried out with the other substrates of Examples 1 and 2.

Example 5

wird zu 2 ml eines leukozytenhaltigen Harnes in einem Reagenzglas gegeben. Der Harn verfärbt sich allmählich - je nach Leukozytenkonzentration - hellgrün bis tiefblau.

is added to 2 ml of urine containing leukocytes in a test tube. The urine gradually changes color - depending on the concentration of leukocytes - from light green to deep blue.

After standing for 10 minutes at room temperature, the leukocyte concentration is determined visually with the aid of reference colors or photometrically, e.g. in 1 cm micro cuvettes at 620 nm.

The sensitivity of the test is around 200 leukocytes / µl urine. The reaction time can be shortened significantly if the incubation is carried out at 37 ° C.

Similar sensitivities (200-1000 leukocytes / µl) can be achieved with the other substrates of Examples 1 and 2, with the addition of organic solvents, e.g. Methanol or dimethylformamide is recommended.

Example 6

Filter paper (eg Schleicher + Schüll 23 SL) is successively impregnated with the following solutions and then dried at 60 ° C.

A colorless test paper is obtained which turns blue when immersed in aqueous solutions which contain the proteolytic enzyme chymotrypsin. Concentrations of 0.02 U chymotrypsin per ml can be detected in about 6-7 minutes.

(The stated enzyme activity was determined with N-acetyl-L-tyrosine ethyl ester as substrate at 25 ° C., pH 7.0 and λ = 237 nm.)

Depending on the amino acid or peptide residue, chymotrypsin or other proteolytic enzymes, such as e.g. Elastase or trypsin, in purely aqueous solutions or also e.g. in body fluids such as Detect whole blood, serum, cerebrospinal fluid, pancreatic secretions or watery stool extracts.

Example 7 3- (N- (toluene-4'-sulfonyl) -L-alanyloxy] indole Solution 1

To prepare the acid chloride using the one-step method, 5.35 g (0.022 mol) of N- (toluene-4-sulfonyl) -L-alanine in 20 ml of abs. Dimethylformamide (DMF) dissolved and cooled to -30 ° C. Then 1.76 ml (0.024 mol) of thionyl chloride are pipetted in with stirring and cooling and the reaction mixture is left in the cold bath at -30 ° C. for 30 minutes.

Solution 2

In a 250 ml three-necked flask equipped with a stirrer, thermometer and a gas inlet and outlet, the solution of 2.90 g (0.022 mol) of indoxyl (3-hydroxy) is added after the air has been completely removed by a gentle stream of nitrogen -indole) in 40 ml abs. DMF, adds 9.74 ml abs. Add pyridine and cool to -15 ° C.

implementation

Solution 1 is poured into solution 2 and stirred with the exclusion of oxygen and water for about 5 h at -15 ° C. until no more indoxyl can be recognized by TLC.

For working up, the reaction mixture is concentrated in vacuo at a maximum bath temperature of 40-50 ° C. The residue is taken up in 100 ml of ethyl acetate and successively twice with 30 ml of 1 N citric acid, 20 ml of water and 50 ml of 5 percent. Washed sodium bicarbonate solution and 25 ml of water. The ethyl acetate phase is - after drying with sodium sulfate - evaporated in vacuo.

The crude product thus obtained is purified by column chromatography on silica gel using a toluene / dioxane mixture (9: 1). After distilling off the solvent of the collected fractions in vacuo, the mixture is obtained by stirring Residue in ether 1.45 g (18.4%) 3- [N- (toluene-4'-sulfonyl) -L-alanlyolxy] indole, colorless crystals, mp. 103 ° C; α20 D: -56.6 °, c = 1% (methanol).

-41.7°, c = 1% (DMF), isolieren.As a by-product, 0.89 g (11.3%) of 1- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -3-hydroxy-indole = 1- [N- (Toluene-4'-sulfonyl) -L-alanyloxy] -3-oxo-indoline, from ethyl acetate: colorless crystals, mp. 187-188 ° C,

Isolate -41.7 °, c = 1% (DMF).

• 7.1. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 123°C,
  
  (Methanol).
• 7.2. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-4-methyl-indol, farblose Kristalle, Schmp. 113 °C,
  
  , c = 1% (Methanol).
• 7.3. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-5-methyl-indol, farblose Kristalle, Schmp. 143-144 °C,
  
  , c = 1% (Methanol).
• 7.4. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-6-methyl-indol, farblose Kristalle, Schmp. 148 °C,
  
  , c = 1 % Methanol).
• 7.5. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-7-methyl-indol, farblose Kristalle, Schmp. 146 °C,
  
  , c = 1 % (Methanol).
• 7.6. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-4,7-dimethyl-indol, farblose Kristalle, Schmp. 121 °C,
  
  , c = 1% (Methanol).
• 7.7. 3-[N-Benzyloxycarbonyl)-L-alanyloxy]-4-chlor-indol, farblose Kristalle, Schmp. 142-143 °C,
  
  , c = 1% (Methanol).
• 7.8. 3-[N-(Benzyloxycarbonyl)-L-alanyloxyl-5-brom-indol, farblose Kristalle, Schmp. 138 °C,
  
  , c = 1% (Methanol).
• 7.9. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-6-chlor-indol, farblose Kristalle, Schmp. 170 °C,
  
  , c = 1% (Methanol).
• 7.10. 3[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4-chlor-5-brom-indol, farblose Kristalle, Schmp. 150-152 °C,
  
  , c = 1 % (Methanol).
• 7.11. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4,5,7-trichlor-indol
• 7.12. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4-chlor-5-brom-7-methyl-indol schwach beigefarbene Kristalle, Schmp. 143-145 °C. DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 2:1, Detektion: UV, NH₃ (Gas), R_F-Wert: 0,56)
• 7.13. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-hydroxy-indol
• 7.14. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-5-methoxy-indol
• 7.15. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-benzyloxy-indol
• 7.16. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4-carboxy-indol
• 7.17. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-4-benzyloxycarbonyl-indol
• 7.18. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-(carboxy-methoxy)-indol
• 7.19. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-(benzyloxy-carbonyl- methoxy)-indol
• 7.20. 3-[N-(Toluol-4'-sulfonyl )-L-alanyloxy]-6-nitro-indol
• 7.21. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-6-acetylamino-indol
• 7.22. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-benzo-[g]-indol
• 7.23. 1-Methyl-3-[N-(benzyloxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 92 °C,
  
  , c = 1% (Methanol).
• 7.24. 1-Benzyl-3-[N-toluol-4'-sulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 149-152 °C,
  
  , c = 1% (DMF).
• 7.25 1-Phenyl-3-[N-(toluol-4'-sulfonyl)-L-alanyloxyl]-indol
• 7.26. 1-Acetyl-3-[N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol
• 7.27. 1-Benzoyl-3-[N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 155-156 °C, α²⁰ _D: -54.7°, c = 1% (DMF)_.
• 7.28. 3-[N-(Benzyloxycarbonyl)-glycyloxy]-indol, farblose Kristalle, Schmp. 122-124 °C.
• 7.29. 3-[N-(Toluol-4'-sulfonyl)-glycyloxy]-indol, farblose Kristalle, Schmp. 103-105 °C.
• 7.30. 3-[N-(Toluol-2'-sulfonyl)-L-alanyloxy]-indol farbloses amorphes Pulver; α²⁰ _D : -62.5°, c = 1 % (Methanol) DC: Fertigplatte Kieselgel (Laufmittel: Toluol-Dioxan 9:1
• Detektion: UV, NH₃ (Gas), R_F-Wert: 0,20) 7.31. 3-[N-(Toluol-3'-sulfonyl)-L-alanyloxy]-indol farblose Kristalle, Schmp. 94°C; α²⁰ _D : - 54.9°, c = 1 % (Methanol)
• 7.32. 3-[N-(Toluol-4'-sulfonyl)-D-alanyloxy]-indol, farblose Kristalle, Schmp. 98 °C, α²⁰ _D +53.1°, c = 1% (Methanol).
• 7.33. 3-[N-(Benzyloxycarbonyl)-D,L-alanyloxy]-indol, farblose Kristalle, Schmp. 110-111 °C.
• 7.34. 3-[N-(Benzyloxycarbonyl)-L-valyloxy]-indol, farblose Kristalle, Schmp. 64-65 °C, α²⁰ _D-41.1°, c = 1% (Methanol).
• 7.35. 3-[N-(Benzyloxycarbonyl)-L-leucyloxyl-indol, farblose Kristalle, Schmp. 73-75 °C, α²⁰D -42.2°, c = 1% (Methanol).
• 7.36. 3-[N-(Benzyloxycarbonyl)-L-isoleucyloxy]-indol, farbloses, amorphes Pulver, α²⁰ _D -30.6°, c = 1 % (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 2:1, Detektion: UV, NH₃ (Gas), R_F-Wert: 0.57).
• 7.37. 3-[N-(Benzyloxycarbonyl)-L-phenylalanyloxy]-indol, gelbliche Kristalle, Schmp. 125 °C, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 9:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.56).
• 7.38. 3-[N-(Toluol-4'-sulfonyl)-L-phenylalanyloxy]-indol, farblose Kristalle, Schmp. 167-169 °C, α²⁰ _D : -34.4°, c = 1% (Methanol).
• 7.39. 3-[N-Acetyl-L-tyrosyloxy]-indol
• 7.40. 3-[N-Benzoyl-L-tyrosyloxy]-indol
• 7.41. 3-[N-(Benzyloxycarbonyl)-L-tyrosyloxy]-indol, farbloser, amorpher Schaum, α²⁰ _D: -22.6°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 4:1, Detektion: UV, NH₃ (Gas), Rr-Wert: 0.27).
• 7.42. 3-[N-(Toluol-4'-sulfonyl)-L-tyrosyloxy]-indol, farblose Kristalle, Schmp. 171 °C, α²⁰ _D: -28.3°, c = 1% (Methanol).
• 7.43. 3-[N-(Toluol-4'-sulfonyl)- O-acetyl-L-tyrosyloxy]-indol, farblose Kristalle, Schmp. 168-170 °C, α²⁰ _D : - 24.1°, c = 1 % (Dimethylformamid).
• 7.44. 3-[N-(Benzyloxycarbonyl)-L-alanyl-L-alanyloxy]-indol, farblose Kristalle, Schmp. 144 °C, α²⁰ _D : -17.3°, c = 1% (Methanol).
• 7.45. 3-[N-(Toluol-4'-sulfonyl)-D-alanyl-L-alanyloxy]-indol, gelblicher, amorpher Schaum, α²⁰ _D : +5.8°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 4:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.19).
• 7.46. 3-[N-(Benzyloxycarbonyl)-L-alanyl-L-alanyl-L-alanyloxy]-indol, farblose Kristalle, Schmp. 156 °C, α²⁰ _D : -39.5°, c = 1% (Methanol).
• 7.47. 3-[N-(Toluol-4'-sulfonyl)-D-alanyl-D-alanyl-L-alanyloxy]-indol, farblose Kristalle, Schmp. 216 °C, α²⁰ _D : +53.8°, c = 1 % (Methanol).
• 7.48. 3-[N-Acetyl-L-alanyloxy]-indol, farbloser, amorpher Schaum, α²⁰ _D: -12.5°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Essigester-Dichlormethan 10:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.33).
• 7.49. 3-[N-Succinyl-L-alanyloxy]-indol, farblose Kristalle, Schmp. 142 °C, α²⁰ _D : - 63.9°, c = 1 % (Methanol).
• 7.50. 3-[N-Benzoyl-D,L-alanyloxy]-indol, farblose Kristalle, Schmp. 171 °C.
• 7.51. 3-[N-Phthaloyl-L-alanyloxy]-indol, farblose Kristalle, Schmp. 58 °C, α²⁰ _D : -26.3°, c = 1% (Methanol).
• 7.52. 3-[N-(Ethoxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 96 °C, α²⁰ _D : -68.9°, c = 1% (Methanol).
• 7.53. 3-[N-(Cyclohexyloxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 149 °C, α²⁰ _D : - 60.4°, c = 1 % (Methanol).
• 7.54. 3-[N-(Phenyloxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 161 °C, α²⁰ _D : -96.0°, c = 1 % (Methanol).
• 7.55. 3-[N-(4'-Methyl-benzyloxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 135 °C, α²⁰ _D : -45.9°, c = 1% (Methanol).
• 7.56. 3-[N-(4'-Nitro-benzyloxycarbonyl)-L-alanyloxy]-indol, gelbliche Kristalle, Schmp. 160 °C, α²⁰ _D : -29.4°, c = 1% (Methanol).
• 7.57. 3-[N-(Benzylthiocarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 122 °C, α²⁰ _D: -77.8°, c = 1% (Methanol).
• 7.58. 3-[N-(Methansulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 164-166 °C, α²⁰ _D : -54.0°, c = 1% (Methanol).
• 7.59. 3-[N-(Benzylsulfonyl)-L-alanyloxy]-indol, farbloses, viskoses Öl, α²⁰ _D : - 54.8°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Essigester 2:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.40).
• 7.60. 3-[N-(Benzolsulfonyl)-L-alanyloxy]-indol, farbloses, viskoses Öl, α²⁰ _D : -59.8°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Essigester 2:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.48).
• 7.61. 3[N-(4'-Brom-benzolsulfonyl)-L-alanyloxy]-indol, gelbliches, amorphes Pulver, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 4:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.39).
• 7.62. 3-[N-(4'-Nitro-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 136-137 °C, α²⁰ _D : -35.9°, c = 1% (Methanol).
• 7.63. 3-[N-(4'-Acetylamino-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 162-163 °C,
  
  , c = 1% (Methanol).
• 7.64. 3-[N-(4'-n-Butyl-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 105 °C,
  
  : -46.1°, c = 1% (Methanol).
• 7.65. 3-[N-(4'-tert.-Butyl-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 140 °C,
  
  , c = 1% (Methanol).
• 7.66. 3-[N-(4'-n-Octyl-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 57 °C,
  
  , c = 1% (Methanol).
• 7.67. 3-[N-(4'-Hydroxy-benzolsulfonyl)-L-alanyloxy]-indol, viskoses, schwach rötliches Öl, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 1:1, Detektion: UV, NH₃ (Gas), R_F-Wert: 0.62).
• 7.68. 3-[N-(4'-Methoxy-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 126-128 °C,
  
  , c = 1% (Methanol).
• 7.69. - 3-[N-(4'-Benzyloxy-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 112°C,
  
  , c = 1% (Methanol).
• 7.70. 3-[N-(4'-(2"-Hydroxy-ethoxy)-benzolsulfonyl)-L-alanyloxy]-indol
• 7.71. 3-[N-(4'-(3"-Oxa-5"-hydroxy-n-pentyloxy)-benzolsulfonyl)-L-alanyloxy]-indol farbloses, viskoses Öl,
  
  , c = 1% (Methanol) DC: Fertigplatte Kieselgel (Laufmittel: Toluol-Essigsäureethylester 1:10, Detektion: UV, NH₃ (Gas), RF-Wert: 0.37
• 7.72. 3-[N-(4'-(3",6"-Di-oxa-n-heptyloxy)-benzolsulfonyl)-L-alanyloxy]-indol, schwach rötliches, viskoses Öl, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 4:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.21
• 7.73. 3-[N-(4'-(2"-Hydroxy-ethyl)-benzolsulfonyl)-L-alanyloxy]-indol, gelbliches, amorphes Pulver,
  
  , c = 1% (Methanol) DC: Fertigplatte, Kieselgel (Laufmittel: Toluol-Essigsäureethylester 1:2, Detektion: UV, NH₃ (Gas), Rr-Wert: 0.26).
• 7.74. 3-[N-(4'-(2"-{4"'-Nitro- benzyloxy}-ethyl)-benzolsulfonyl)-L-alanyloxy]-indol
• 7.75. 3-[N-(4'-(2"-Chlor-ethyl)-benzolsulfonyl)-L-alanyloxy]-indol, schwach rötlicher, amorpher Schaum,
  
  , c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Essigester 5:1, Detektion: UV, NH3 (Gas), R_F-Wert: 0.20).
• 7.76. 3-[N-(4'-Cyano-benzolsulfonyl)-L-alanyloxy]-indol farblose Kristalle, Schmp. 131 °C;
  
  , c = 1% (Dimethylformamid).
• 7.77. 3-[N-(4'-Carboxy-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 143 °C,
  
  , c = 1% (Methanol).
• 7.78. 3-[N-(4'-Benzyloxycarbonyl- benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 178 °C,
  
  , c = 1 % (Aceton).
• 7.79. 3-[N-(4'-Carbamoyl-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 206 °C,
  
  , c = 1% (Aceton).
• 7.80. 3-[N-(4'-(Dimethyl-carbamoyl)-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 170-172 °C,
  
  , c = 1 % (Dimethylformamid).
• 7.81. 3-[N-Methyl-N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 82 °C,
  
  , c = 1% (Methanol).
• 7.82. 3-[N-(2',4',6'-Trimethyl- benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 192-194 °C,
  
  , c = 1 % (Methanol).
• 7.83. 3-[N-(Biphenyl-4'-sulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 159°C,
  
  , c = 1% (Methanol).
• 7.84. 3-[N-(Naphthalin-2'-sulfonyl)-L-alanyloxy]-indol, schwach rötliche Kristalle, Schmp. 103-105 °C, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 4:1, Detektion: UV, NH₃ (Gas), R_F-Wert: 0.41).
• 7.85. 3-[N-(4'-Acetylamino-naphthalin-1'-sulfonyl)-Lalanyloxy]-indol, farblose Kristalle, Schmp. 129-132 °C,
  
  , c = 1 % (Methanol).
• 7.86. 3-[N-(5'-Dimethylamino-naphthalin-1'-sulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 170 °C,
  
  , c = 1 % (Methanol).
• 7.87. 3-[N-(Benzyloxycarbonyl)-L-alanyloxy]-benzo-[b]-thiophen, farbloses, viskoses Öl,
  
  , c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Dichlormethan, Detektion: UV, NaOH / K₃Fe(CN)₆, R_F-Wert: 0.13).
• 7.88. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-benzyl-indol
• 7.89. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-phenyl-indol

In an analogous manner, the following substrates are obtained by reacting the appropriately substituted indoxyl compounds with the respective amino acids, with the above-mentioned 1-substituted 3-hydroxyindoles being formed as by-products in all cases (except 7.23. - 7.27.):

• 7.1. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 123 ° C,
  
  (Methanol).
• 7.2. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -4-methyl-indole, colorless crystals, mp. 113 ° C.,
  
  , c = 1% (methanol).
• 7.3. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -5-methyl-indole, colorless crystals, mp. 143-144 ° C,
  
  , c = 1% (methanol).
• 7.4. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -6-methyl-indole, colorless crystals, mp. 148 ° C,
  
  , c = 1% methanol).
• 7.5. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -7-methyl-indole, colorless crystals, mp. 146 ° C,
  
  , c = 1% (methanol).
• 7.6. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -4,7-dimethyl-indole, colorless crystals, mp. 121 ° C.,
  
  , c = 1% (methanol).
• 7.7. 3- [N-Benzyloxycarbonyl) -L-alanyloxy] -4-chloroindole, colorless crystals, mp. 142-143 ° C,
  
  , c = 1% (methanol).
• 7.8. 3- [N- (Benzyloxycarbonyl) -L-alanyloxyl-5-bromo-indole, colorless crystals, mp. 138 ° C.,
  
  , c = 1% (methanol).
• 7.9. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -6-chloroindole, colorless crystals, mp. 170 ° C,
  
  , c = 1% (methanol).
• 7.10. 3 [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-chloro-5-bromo-indole, colorless crystals, mp. 150-152 ° C,
  
  , c = 1% (methanol).
• 7.11. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4,5,7-trichloroindole
• December 7th 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-chloro-5-bromo-7-methyl-indole slightly beige crystals, mp. 143-145 ° C. TLC: finished plate silica gel, (eluent: toluene-dioxane 2: 1, detection: UV, NH ₃ (gas), R _F value: 0.56)
• 7.13. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-hydroxy-indole
• 7.14. 3- [N- (Benzyloxycarbonyl) -L-alanyloxy] -5-methoxy-indole
• 7.15. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-benzyloxy-indole
• 7.16. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-carboxy-indole
• 7.17. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -4-benzyloxycarbonyl-indole
• 7.18. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5- (carboxymethoxy) indole
• 7.19. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5- (benzyloxy-carbonyl-methoxy) indole
• 7.20. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -6-nitro-indole
• 7.21. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -6-acetylamino-indole
• 7.22. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] benzo [g] indole
• 7.23. 1-methyl-3- [N- (benzyloxycarbonyl) -L-alanyloxy] indole, colorless crystals, melting point 92 ° C.,
  
  , c = 1% (methanol).
• 7.24. 1-benzyl-3- [N-toluene-4'-sulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 149-152 ° C,
  
  , c = 1% (DMF).
• 7.25 1-phenyl-3- [N- (toluene-4'-sulfonyl) -L-alanyloxyl] indole
• 7.26. 1-acetyl-3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] indole
• 7.27. 1-Benzoyl-3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] indole, colorless crystals, mp 155-156 ° C, α ²⁰ _D : -54.7 °, c = 1% (DMF ) _.
• 7.28. 3- [N- (Benzyloxycarbonyl) glycyloxy] indole, colorless crystals, mp 122-124 ° C.
• 7.29. 3- [N- (Toluene-4'-sulfonyl) glycyloxy] indole, colorless crystals, mp 103-105 ° C.
• 7.30. 3- [N- (toluene-2'-sulfonyl) -L-alanyloxy] indole colorless amorphous powder; α ²⁰ _D : -62.5 °, c = 1% (methanol) TLC: silica gel finished plate (eluent: toluene-dioxane 9: 1
• Detection: UV, NH ₃ (gas), R _F value: 0.20) 7.31. 3- [N- (toluene-3'-sulfonyl) -L-alanyloxy] indole colorless crystals, mp 94 ° C; α ²⁰ _D : - 54.9 °, c = 1% (methanol)
• 7.32. 3- [N- (Toluene-4'-sulfonyl) -D-alanyloxy] indole, colorless crystals, mp. 98 ° C, α ²⁰ _D + 53.1 °, c = 1% (methanol).
• 7.33. 3- [N- (Benzyloxycarbonyl) -D, L-alanyloxy] indole, colorless crystals, mp 110-111 ° C.
• 7.34. 3- [N- (Benzyloxycarbonyl) -L-valyloxy] indole, colorless crystals, mp. 64-65 ° C, α ²⁰ _D -41.1 °, c = 1% (methanol).
• 7.35. 3- [N- (Benzyloxycarbonyl) -L-leucyloxyl-indole, colorless crystals, mp. 73-75 ° C, α ²⁰ D -42.2 °, c = 1% (methanol).
• 7.36. 3- [N- (Benzyloxycarbonyl) -L-isoleucyloxy] indole, colorless, amorphous powder, α ²⁰ _D -30.6 °, c = 1% (methanol), TLC: finished plate silica gel, (eluent: toluene-dioxane 2: 1 , Detection: UV, NH ₃ (gas), R _F value: 0.57).
• 7.37. 3- [N- (Benzyloxycarbonyl) -L-phenylalanyloxy] indole, yellowish crystals, mp. 125 ° C, TLC: silica gel finished plate, (eluent: toluene-dioxane 9: 1, detection: UV, NH ₃ (gas), RF value: 0.56).
• 7.38. 3- [N- (toluene-4'-sulfonyl) -L-phenylalanyloxy] indole, colorless crystals, mp. 167-169 ° C, α ²⁰ _D : -34.4 °, c = 1% (methanol).
• 7.39. 3- [N-acetyl-L-tyrosyloxy] indole
• 7.40. 3- [N-benzoyl-L-tyrosyloxy] indole
• 7.41. 3- [N- (Benzyloxycarbonyl) -L-tyrosyloxy] indole, colorless, amorphous foam, α ²⁰ _D : -22.6 °, c = 1% (methanol), TLC: finished plate silica gel, (eluent: toluene-dioxane 4: 1, detection: UV, NH ₃ (gas), Rr value: 0.27).
• 7.42. 3- [N- (Toluene-4'-sulfonyl) -L-tyrosyloxy] indole, colorless crystals, mp. 171 ° C, α ²⁰ _D : -28.3 °, c = 1% (methanol).
• 7.43. 3- [N- (toluene-4'-sulfonyl) - O-acetyl-L-tyrosyloxy] indole, colorless crystals, mp. 168-170 ° C, α ²⁰ _D : - 24.1 °, c = 1% (dimethylformamide ).
• 7.44. 3- [N- (Benzyloxycarbonyl) -L-alanyl-L-alanyloxy] indole, colorless crystals, mp. 144 ° C, α ²⁰ _D : -17.3 °, c = 1% (methanol).
• 7.45. 3- [N- (Toluene-4'-sulfonyl) -D-alanyl-L-alanyloxy] indole, yellowish, amorphous foam, α ²⁰ _D : + 5.8 °, c = 1% (methanol), TLC: finished plate silica gel , (Eluent: toluene-dioxane 4: 1, detection: UV, NH ₃ (gas), RF value: 0.19).
• 7.46. 3- [N- (Benzyloxycarbonyl) -L-alanyl-L-alanyl-L-alanyloxy] indole, colorless crystals, mp. 156 ° C, α ²⁰ _D : -39.5 °, c = 1% (methanol).
• 7.47. 3- [N- (Toluene-4'-sulfonyl) -D-alanyl-D-alanyl-L-alanyloxy] indole, colorless crystals, mp. 216 ° C, α ²⁰ _D : + 53.8 °, c = 1% (Methanol).
• 7.48. 3- [N-acetyl-L-alanyloxy] indole, colorless, amorphous foam, α ²⁰ _D : -12.5 °, c = 1% (methanol), TLC: finished plate silica gel, (mobile phase: ethyl acetate-dichloromethane 10: 1, Detection: UV, NH ₃ (gas), RF value: 0.33).
• 7.49. 3- [N-succinyl-L-alanyloxy] indole, colorless crystals, mp. 142 ° C, α ²⁰ _D : - 63.9 °, c = 1% (methanol).
• 7.50. 3- [N-Benzoyl-D, L-alanyloxy] indole, colorless crystals, mp. 171 ° C.
• 7.51. 3- [N-phthaloyl-L-alanyloxy] indole, colorless crystals, mp. 58 ° C, α ²⁰ _D : -26.3 °, c = 1% (methanol).
• 7.52. 3- [N- (Ethoxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 96 ° C, α ²⁰ _D : -68.9 °, c = 1% (methanol).
• 7.53. 3- [N- (Cyclohexyloxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 149 ° C, α ²⁰ _D : - 60.4 °, c = 1% (methanol).
• 7.54. 3- [N- (Phenyloxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 161 ° C, α ²⁰ _D : -96.0 °, c = 1% (methanol).
• 7.55. 3- [N- (4'-Methyl-benzyloxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 135 ° C, α ²⁰ _D : -45.9 °, c = 1% (methanol).
• 7.56. 3- [N- (4'-Nitro-benzyloxycarbonyl) -L-alanyloxy] indole, yellowish crystals, mp. 160 ° C, α ²⁰ _D : -29.4 °, c = 1% (methanol).
• 7.57. 3- [N- (Benzylthiocarbonyl) -L-alanyloxy] indole, colorless crystals, mp 122 ° C, α ²⁰ _D : -77.8 °, c = 1% (methanol).
• 7.58. 3- [N- (methanesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 164-166 ° C, α ²⁰ _D : -54.0 °, c = 1% (methanol).
• 7.59. 3- [N- (Benzylsulfonyl) -L-alanyloxy] indole, colorless, viscous oil, α ²⁰ _D : - 54.8 °, c = 1% (methanol), TLC: finished plate silica gel, (mobile phase: toluene-ethyl acetate 2: 1, detection: UV, NH ₃ (gas), RF value: 0.40).
• 7.60. 3- [N- (benzenesulfonyl) -L-alanyloxy] indole, colorless, viscous oil, α ²⁰ _D : -59.8 °, c = 1% (methanol), TLC: finished plate silica gel, (eluent: toluene-ethyl acetate 2: 1, detection: UV, NH ₃ (gas), RF value: 0.48).
• 7.61. 3 [N- (4'-bromo-benzenesulfonyl) -L-alanyloxy] indole, yellowish, amorphous powder, TLC: silica gel finished plate, (mobile phase: toluene-dioxane 4: 1, Detection: UV, NH ₃ (gas), RF value: 0.39).
• 7.62. 3- [N- (4'-Nitro-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp 136-137 ° C, α ²⁰ _D : -35.9 °, c = 1% (methanol).
• 7.63. 3- [N- (4'-Acetylamino-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp 162-163 ° C,
  
  , c = 1% (methanol).
• 7.64. 3- [N- (4'-n-Butyl-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 105 ° C.,
  
  : -46.1 °, c = 1% (methanol).
• 7.65. 3- [N- (4'-tert-butyl-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 140 ° C.,
  
  , c = 1% (methanol).
• 7.66. 3- [N- (4'-n-octyl-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 57 ° C.,
  
  , c = 1% (methanol).
• 7.67. 3- [N- (4'-Hydroxy-benzenesulfonyl) -L-alanyloxy] indole, viscous, slightly reddish oil, TLC: finished plate silica gel, (eluent: toluene-dioxane 1: 1, detection: UV, NH ₃ (gas ), R _F value: 0.62).
• 7.68. 3- [N- (4'-methoxy-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 126-128 ° C,
  
  , c = 1% (methanol).
• 7.69. 3- [N- (4'-benzyloxy-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp 112 ° C.,
  
  , c = 1% (methanol).
• 7.70. 3- [N- (4 '- (2 "-hydroxy-ethoxy) -benzenesulfonyl) -L-alanyloxy] indole
• 7.71. 3- [N- (4 '- (3 "-Oxa-5" -hydroxy-n-pentyloxy) benzenesulfonyl) -L-alanyloxy] indole colorless, viscous oil,
  
  , c = 1% (methanol) TLC: silica gel finished plate (eluent: toluene-ethyl acetate 1:10, detection: UV, NH ₃ (gas), RF value: 0.37
• 7.72. 3- [N- (4 '- (3 ", 6" -Di-oxa-n-heptyloxy) -benzenesulfonyl) -L-alanyloxy] -indole, slightly reddish, viscous oil, TLC: finished plate silica gel, (eluent: toluene -Dioxane 4: 1, detection: UV, NH ₃ (gas), RF value: 0.21
• 7.73. 3- [N- (4 '- (2 "-hydroxy-ethyl) -benzenesulfonyl) -L-alanyloxy] indole, yellowish, amorphous powder,
  
  , c = 1% (methanol) TLC: finished plate, silica gel (mobile phase: toluene-ethyl acetate 1: 2, detection: UV, NH ₃ (gas), Rr value: 0.26).
• 7.74. 3- [N- (4 '- (2 "- {4"' - nitrobenzyloxy} ethyl) benzenesulfonyl) -L-alanyloxy] indole
• 7.75. 3- [N- (4 '- (2 "-chloro-ethyl) -benzenesulfonyl) -L-alanyloxy] indole, slightly reddish, amorphous foam,
  
  , c = 1% (methanol), TLC: silica gel finished plate, (mobile phase: toluene-ethyl acetate 5: 1, detection: UV, NH3 (gas), R _F value: 0.20).
• 7.76. 3- [N- (4'-Cyano-benzenesulfonyl) -L-alanyloxy] indole colorless crystals, mp 131 ° C;
  
  , c = 1% (dimethylformamide).
• 7.77. 3- [N- (4'-carboxy-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 143 ° C.,
  
  , c = 1% (methanol).
• 7.78. 3- [N- (4'-Benzyloxycarbonylbenzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 178 ° C,
  
  , c = 1% (acetone).
• 7.79. 3- [N- (4'-Carbamoyl-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 206 ° C.,
  
  , c = 1% (acetone).
• 7.80. 3- [N- (4 '- (Dimethyl-carbamoyl) -benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 170-172 ° C,
  
  , c = 1% (dimethylformamide).
• 7.81. 3- [N-methyl-N- (toluene-4'-sulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 82 ° C.,
  
  , c = 1% (methanol).
• 7.82. 3- [N- (2 ', 4', 6'-trimethylbenzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 192-194 ° C,
  
  , c = 1% (methanol).
• 7.83. 3- [N- (Biphenyl-4'-sulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 159 ° C.,
  
  , c = 1% (methanol).
• 7.84. 3- [N- (naphthalene-2'-sulfonyl) -L-alanyloxy] indole, slightly reddish crystals, mp. 103-105 ° C, TLC: finished plate silica gel, (eluent: toluene-dioxane 4: 1, detection: UV, NH ₃ (gas), R _F value: 0.41).
• 7.85. 3- [N- (4'-acetylamino-naphthalene-1'-sulfonyl) -lalanyloxy] indole, colorless crystals, mp. 129-132 ° C,
  
  , c = 1% (methanol).
• 7.86. 3- [N- (5'-Dimethylamino-naphthalene-1'-sulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 170 ° C.,
  
  , c = 1% (methanol).
• 7.87. 3- [N- (benzyloxycarbonyl) -L-alanyloxy] benzo [b] thiophene, colorless, viscous oil,
  
  , c = 1% (methanol), TLC: silica gel finished plate, (eluent: dichloromethane, detection: UV, NaOH / K ₃ Fe (CN) ₆ , R _F value: 0.13).
• 7.88. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-benzyl-indole
• 7.89. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-phenyl-indole

Example 8 3- [N- (tert-Butyloxycarbonyl) -L-alanyloxy] indole

For the reaction according to the carbodimide process, 3.23 g (0.017 mol) of N-tert-butyloxycarbonyl-L-alanine and 2.27 g (0.017 mol) of indoxyl in 100 ml of abs. Dioxane-dichloromethane dissolved 1: 1. After adding a solution of 3.87 g (0.019 mol) of dicyclohexylcarbodiimide (DCC) in 20 ml of abs. Dioxane is stirred with the exclusion of water and oxygen for 68 h at room temperature. The deposited N, N'-dicyclohexylurea is filtered off with suction, the solvent is distilled off in vacuo and the residue is taken up in about 100 ml of ethyl acetate. After suctioning off further precipitated N, N'-dicyclohexylurea, the clear ethyl acetate solution is in succession twice with 30 ml of 1 N citric acid, 20 ml of water, 50 ml of 5-10 percent. Washed sodium bicarbonate solution and 25 ml of water. After drying over sodium sulfate, the ethyl acetate phase is concentrated in vacuo. The amorphous residue is purified by column chromatography on silica gel using a toluene / dioxane mixture (9: 1).

• 1.04 g (20%) 3-[N-(tert. -Butyloxycarbonyl)-L-alanyloxy]-indol,
• farblose Kristalle, Schmp. 158 °C,
• 
  , c = 1% (Methanol).

After concentrating the collected fractions in vacuo and recrystallizing the residue from ethyl acetate-ether (1:10)

• 1.04 g (20%) 3- [N- (tert -Butyloxycarbonyl) -L-alanyloxy] indole,
• colorless crystals, mp. 158 ° C,
• 
  , c = 1% (methanol).

This reaction also produces the 3-hydroxy-indole substituted in the 1-position as a by-product (see Example 7).

• 8.1. 3-[N-Formyl-L-alanyloxy]-indol, farblose Kristalle, Schmp. 121-122 °C,
  
  , c = 1 % (Methanol).
• 8.2. 3-[N-(4 '-Methoxy-benzyloxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 92 °C,
  
  , c = 1% (Methanol).
• 8.3. 3-[N-(N-(Piperidino)-oxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 104 °C,
  
  , c = 1% (Methanol).
• 8.4. 3-[N-(Thienyl-(2')-methoxycarbonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 134 °C,
  
  , c = 1 % (Methanol).
• 8.5. 3-[N-(Chinolin-8'-sulfonyl)-L-alanyloxy]-indol, amorphes, schwach rötliches Pulver, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 4:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.23).
• 8.6. 3-[N-(Diphenylcarbamoyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 188-190 °C, α²⁰ _D : +22.2°, c = 1% (Pyridin).

The following substrates are obtained in an analogous manner by reacting the correspondingly substituted indoxyl compounds with the respective amino acids, the corresponding 3-hydroxyindoles substituted in the 1-position also being formed here in all cases:

• 8.1. 3- [N-formyl-L-alanyloxy] indole, colorless crystals, mp. 121-122 ° C,
  
  , c = 1% (methanol).
• 8.2. 3- [N- (4 '-Methoxy-benzyloxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 92 ° C,
  
  , c = 1% (methanol).
• 8.3. 3- [N- (N- (Piperidino) -oxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 104 ° C,
  
  , c = 1% (methanol).
• 8.4. 3- [N- (Thienyl- (2 ') - methoxycarbonyl) -L-alanyloxy] indole, colorless crystals, mp. 134 ° C,
  
  , c = 1% (methanol).
• 8.5. 3- [N- (Quinoline-8'-sulfonyl) -L-alanyloxy] indole, amorphous, slightly reddish powder, TLC: silica gel finished plate, (eluent: toluene-dioxane 4: 1, detection: UV, NH ₃ (gas ), RF value: 0.23).
• 8.6. 3- [N- (Diphenylcarbamoyl) -L-alanyloxy] indole, colorless crystals, mp. 188-190 ° C, α ²⁰ _D : + 22.2 °, c = 1% (pyridine).

Example 9 3- [N- (furyl- (2 ') methoxycarbonyl) -L-alanyloxy] indole

• 3.0 g (45.7%) 3-[N-(Furyl-(2')-methoxycarbonyl)-L-alanyloxy]-indol,
• farblose Kristalle, Schmp. 129 °C,
• 
  , c = 1 % (Methanol).

For the reaction according to the active ester method, 4.26 g (0.02 mol) of N- (furyl- (2 ') - methoxycarbonyl) -L-alanine and 5.4 g (0.04 mol) of N-hydroxy-benzotriazole in 50 ml abs. Dissolved tetrahydrofuran (THF), cooled to 0 ° C and with a solution of 4.4 g (0.022 mol) of dicyclohexylcarbodiimide (DCC) in 10 ml abs. THF offset. To form the active ester, the reaction mixture is stirred at 0 ° C. for 1.5 h, then at room temperature for 2 h. Then add 2.66 g (0.02 mol) indoxyl and 2.77 ml (0.04 mol) abs while excluding oxygen and water. Triethylamine too. The mixture is stirred at room temperature for 18 h. The N, N'-dicyclohexylurea formed is filtered off with suction, the solvent is distilled off in vacuo and the residue is worked up as described in Example 7. The amorphous crude product is then purified by column chromatography on a silica gel column using a toluene / ethyl acetate mixture (5: 1). After concentration of the corresponding collected fractions and recrystallization from ethyl acetate-petroleum ether 1: 1

• 3.0 g (45.7%) 3- [N- (furyl- (2 ') methoxycarbonyl) -L-alanyloxy] indole,
• colorless crystals, mp. 129 ° C,
• 
  , c = 1% (methanol).

• 9.1. 3-[N-(Toluol-4'-sulfonyl-L-valyloxy]-indol, farblose Kristalle, Schmp. 125-127 °C,
  
  , c = 1% (Methanol).
• 9.2. 3-[N-(4'-Dimethylamino-benzolsulfonyl)-L-alanyloxy]-indol, amorphes, schwach rötliches Pulver, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 3:1, Detektion: UV, NH₃ (Gas), RF-Wert: 0.25).
• 9. 3. 3-[N-(4'-Acetyl-benzolsulfonyl)-L-alanyloxy]-indol
• 9.4. 3-[N-(4'-Methoxycarbonyl-benzolsulfo- nyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 140-141 °C,
  
  , c = 1 % (Methanol).
• 9.5. 3-[N-(4'-Carboxymethyl-benzolsulfonyl)-L-alanyloxy]-indol, amorpher, farbloser Schaum, α²⁰ _D : -82.1°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Isopropanol-Essigsäure-n-butylester-Wasser 5:3:2, Detektion: UV, NH₃ (Gas), R_FWert: 0.68).
• 9.6. 3-[N-(4'Carboxymethoxy-benzolsulfonyl)-L-alanyloxy]-indol, amorpher, farbloser Schaum, α²⁰ _D : -47.7°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan-Eisessig 6:2:1, Detektion: UV, NH₃ (Gas), RFWert: 0.27).
• 9.7. 3-[N-(4'-Carboxymethylamino- benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 151-153 °C α²⁰ _D : -71.5°, c = 1% (Methanol).
• 9.8. 3-[N'-(4'-(Benzyloxycarbonyl- methylamino)-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 110-113 °C, α²⁰ _D : - 60.0°, c = 1 % (Methanol).
• 9.9. 3-[N-(4'-Fluor-benzolsulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 94-95 °C, α²⁰ _D : -48.5°, c = 1% (Methanol).
• 9.10. 3-[N-(4'-Fluorsulfonyl-benzolsulfonyl)-L-alanyloxy]-indol
• 9.11. 3-[N-(4'-Sulfamoyl-benzolsulfonyl)-L-alanyloxy]-indol
• 9.12. 3-[N-Acetyl-N-(toluol-4'-sulfonyl)-L-alanyloxy]-indol, farbloses, amorphes Pulver, α²⁰ _D : -10.6°, c = 1% (Methanol), DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 9:1, Detektion: UV, NH₃ (Gas), R_FWert: 0.17).
• 9.13. 3-[N-(Pyridin-3'- sulfonyl)-L-alanyloxy]-indol, farblose Kristalle, Schmp. 155-157 °C, α²⁰ _D : -55.1°, c = 1% (Methanol).
• 9.14. 3-[N-(5',5'-Dimethyl-3'-oxo- cyclohexen-1'-yl)-L-alanyloxy]-indol. farblose Kristalle, Schmp. 154°C, α²⁰ _D : -272.2°, c = 1% (Methanol).
• 9.15. 3-[N-(Di-(4'-nitrobenzyl)-phosphoryl)-L-alanyloxy]-indol
• 9.16. 3-[N-(Di-(4'-brombenzyl)-phosphoryl)-L-alanyloxy]-indol
• 9.17. 3-[N-(Toluol-4'-sulfonyl)-L-alanyloxy]-5-methoxy-indol farbloses, viskoses Öl; α²⁰ _D : - 46.8°, c = 1% (Methanol), DC:Fertigplatte, Kieselgel (Laufmittel: Toluol-Essigsäureethylester 2:1 Detektion: UV, NH₃ (Gas), R_FWert: 0.45).

The following substances are obtained in an analogous manner by reacting the correspondingly substituted amino acids or peptides with the respective indoxyl compounds:

• 9.1. 3- [N- (toluene-4'-sulfonyl-L-valyloxy] indole, colorless crystals, mp 125-127 ° C,
  
  , c = 1% (methanol).
• 9.2. 3- [N- (4'-Dimethylamino-benzenesulfonyl) -L-alanyloxy] indole, amorphous, slightly reddish powder, TLC: finished plate silica gel, (eluent: toluene-dioxane 3: 1, detection: UV, NH ₃ (gas ), RF value: 0.25).
• 9. 3. 3- [N- (4'-Acetyl-benzenesulfonyl) -L-alanyloxy] indole
• 9.4. 3- [N- (4'-methoxycarbonyl-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 140-141 ° C,
  
  , c = 1% (methanol).
• 9.5. 3- [N- (4'-Carboxymethyl-benzenesulfonyl) -L-alanyloxy] indole, amorphous, colorless foam, α ²⁰ _D : -82.1 °, c = 1% (methanol), TLC: finished plate silica gel, (eluent: Isopropanol-n-butyl ester water 5: 3: 2, detection: UV, NH ₃ (gas), R _F value: 0.68).
• 9.6. 3- [N- (4'Carboxymethoxy-benzenesulfonyl) -L-alanyloxy] indole, amorphous, colorless foam, α ²⁰ _D : -47.7 °, c = 1% (methanol), TLC: silica gel ready plate, (eluent: toluene -Dioxane-glacial acetic acid 6: 2: 1, detection: UV, NH ₃ (gas), RF value: 0.27).
• 9.7. 3- [N- (4'-Carboxymethylamino-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp 151-153 ° C α ²⁰ _D : -71.5 °, c = 1% (methanol).
• 9.8. 3- [N '- (4' - (Benzyloxycarbonyl-methylamino) -benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 110-113 ° C, α ²⁰ _D : - 60.0 °, c = 1% ( Methanol).
• 9.9. 3- [N- (4'-Fluoro-benzenesulfonyl) -L-alanyloxy] indole, colorless crystals, mp. 94-95 ° C, α ²⁰ _D : -48.5 °, c = 1% (methanol).
• 9.10. 3- [N- (4'-Fluorosulfonyl-benzenesulfonyl) -L-alanyloxy] indole
• 9.11. 3- [N- (4'-Sulfamoyl-benzenesulfonyl) -L-alanyloxy] indole
• 9.12. 3- [N-acetyl-N- (toluene-4'-sulfonyl) -L-alanyloxy] indole, colorless, amorphous powder, α ²⁰ _D : -10.6 °, c = 1% (methanol), TLC: finished plate silica gel , (Eluent: toluene-dioxane 9: 1, detection: UV, NH ₃ (gas), R _F value: 0.17).
• 9.13. 3- [N- (pyridine-3'-sulfonyl) -L-alanyloxy] indole, colorless crystals, mp 155-157 ° C, α ²⁰ _D : -55.1 °, c = 1% (methanol).
• 9.14. 3- [N- (5 ', 5'-Dimethyl-3'-oxocyclohexen-1'-yl) -L-alanyloxy] indole. colorless crystals, mp. 154 ° C, α ²⁰ _D : -272.2 °, c = 1% (methanol).
• 9.15. 3- [N- (Di- (4'-nitrobenzyl) phosphoryl) -L-alanyloxy] indole
• 9.16. 3- [N- (Di- (4'-bromobenzyl) phosphoryl) -L-alanyloxy] indole
• 9.17. 3- [N- (toluene-4'-sulfonyl) -L-alanyloxy] -5-methoxy-indole colorless, viscous oil; α ²⁰ _D : - 46.8 °, c = 1% (methanol), TLC: finished plate, silica gel (eluent: toluene-ethyl acetate 2: 1 detection: UV, NH ₃ (gas), R _F value: 0.45).

Example 10 3- [N- (3 ', 6'-Dioxa-n-heptyloxycarbonyl) -L-alanyloxy] indole

• 0.51 g (9.5%) 3-[N-(3',6'-Dioxa-n-heptyloxy- carbonyl)-L alanyloxy]-indol,
• farbloses, viskoses Öl,
• α²⁰ _D : -40.4°, c = 1% (Methanol),
• DC: Fertigplatte Kieselgel,
• (Laufmittel: Toluol-Methylethylketon 1:2,
• Detektion: UV, NH₃ (Gas),
• R_F-Wert: 0.54).

For the reaction according to the mixed anhydride method, 3.52 g (0.015 mol) of N- (3,6-dioxa-n-heptyloxy-carbonyl) -L-alanine in 35 ml abs. Tetrahydrofuran (THF) dissolved. After adding 2.1 ml (0.015 mol) of triethylamine, the mixture is cooled to -15 ° C. and then 1.43 ml (0.015 mol) of ethyl chloroformate are added to form the mixed anhydride. After stirring for 1 h at -15 ° C, a solution of 2.00 g (0.015 mol) indoxyl in 20 ml abs, cooled to -15 ° C, is added with the exclusion of oxygen and water. THF too. The mixture is stirred for a further 2 h at -15 ° C to - 20 ° C and then left in the refrigerator overnight. The triethylamine hydrochloride formed is filtered off, the solvent is removed in vacuo and the residue is worked up as described in Example 7. The amorphous crude product is then purified by column chromatography on silica gel first with a methylene chloride-methanol mixture (95: 5) and then with a toluene-methyl ethyl ketone mixture (1: 2). After treating the collected fractions with activated carbon and removing the solvent

• 0.51 g (9.5%) 3- [N- (3 ', 6'-dioxa-n-heptyloxycarbonyl) -L alanyloxy] indole,
• colorless, viscous oil,
• α ²⁰ _D : -40.4 °, c = 1% (methanol),
• DC: finished plate silica gel,
• (Eluent: toluene-methyl ethyl ketone 1: 2,
• Detection: UV, NH ₃ (gas),
• R _F value: 0.54).

• 10.1. 3-[N-(2'-Nitro-benzolsulfenyl)-L-alanyloxy]-indol, gelbe Kristalle, Schmp. 133-134°C,
  
  , c = 1% (Methanol).
• 10.2. 3-[N-(1'-Methyl-2'-benzoyl- vinyl)-L-alanyloxy]-indol, schwach rötliche Kristalle, Schmp. 130-133 °C, DC: Fertigplatte Kieselgel, (Laufmittel: Toluol-Dioxan 4:1, Detektion: UV, NH₃ (Gas), R_F-Wert: 0.45).

The following substances are obtained in an analogous manner by reaction of the correspondingly substituted amino acids with the respective indoxyl compounds:

• 10.1. 3- [N- (2'-Nitro-benzenesulfenyl) -L-alanyloxy] indole, yellow crystals, mp. 133-134 ° C,
  
  , c = 1% (methanol).
• 10.2. 3- [N- (1'-Methyl-2'-benzoyl-vinyl) -L-alanyloxy] indole, slightly reddish crystals, mp. 130-133 ° C, TLC: finished plate silica gel, (eluent: toluene-dioxane 4 : 1, detection: UV, NH ₃ (gas), R _F value: 0.45).

Claims (6)

1. Diagnostic agent for the detection of proteolytic enzymes, consisting of an absorbent carrier, a film layer, a powder mixture, a lyophilisate, a solution or a reagent tablet, containing one or more chromogens and a suitable buffer substance, characterised in that as chromogens there are used indoxyl- and/or thioindoxylamino acid esters and/or peptide esters of the general formula I

in which

R₁, R₂, R₃, R₄, which can be the same or different, each signify hydrogen or halogen, a lower alkyl, lower alkoxy, aryl, aralkyl, aralkoxy, a hydroxyl, a carboxy, a carboxy lower alkoxy, an aralkoxycarbonyl, an aralkyloxycarbonyl lower alkoxy, a nitro or a lower acylamino group or two neighbouring substituents a benzo-annellated radical possibly substituted by halogen,

X a sulphur atom or an imino group possibly substituted by a lower alkyl, an aryl, an aralkyl or an acyl radical,

A an amino acid or a peptide residue,

B a nitrogen protective group conventional in peptide chemistry or derived therefrom.

2. Diagnostic agent according to claim 1, characterised in that conventionally used adjuvants are additionally added.

3. Diagnostic agent according to claim 2, characterised in that wetting agents, oxidation agents, film formers, galenical additives and/or structural formers are employed as additional ajuvants.

4. Use of indoxyl- and/or thioindoxyl-amino acid esters and/or peptide esters of the general formula I

in which

R₁, R₂, R₃, R₄, which can be the same or different, each signify hydrogen or halogen, a lower alkyl, lower alkoxy, aryl, aralkyl, aralkoxy, a hydroxyl, a carboxy, a carboxy lower alkoxy, an aralkoxycarbonyl, an aralkyloxycarbonyl lower alkoxy, a nitro or a lower acylamino group of two neighbouring substituents a benzo-annellated radical possibly substituted by halogen,

X a sulphur atom or an imino group possibly substituted by a lower alkyl, an aryl, an aralkyl or an acyl radical,

A an amino acid or a peptide residue,

B a nitrogen protective group conventional in peptide chemistry or derived therefrom, for the production of diagnostic agents for the detection of proteolytic enzymes.

5. Indoxyl and thioindoxyl-amino acid esters and peptide esters of the general formula I

in which

R_i, R₂, R₃, R₄, which can be the same or different, each signify hydrogen or halogen, a lower alkyl, lower alkoxy, aryl, aralkyl, aralkoxy, a hydroxyl, a carboxy, a carboxy lower alkoxy, an aralkoxycarbonyl, an aralkyloxycarbonyl lower alkoxy, a nitro or a lower acylamino group or two neighbouring substituents a benzo-annellated radical possibly substituted by halogen,

X a sulphur atom or an imino group possibly substituted by a lower alkyl, an aryl, an aralkyl or an acyl radical,

A an amino acid or a peptide residue,

B a nitrogen protective group conventional in peptide chemistry or derived therefrom.

6. Process for the preparation of indoxyl- and thioindoxyl-amino acid esters and peptide esters of the general formula I

in which

R₁, R₂, R₃, R₄, which can be the same or different, each signify hydrogen or halogen, a lower alkyl, lower alkoxy, aryl, aralkyl, aralkoxy, a hydroxyl, a carboxy, a carboxy lower alkoxy, an aralkoxycarbonyl, an aralkyloxycarbonyl lower alkoxy, a nitro or a lower acylamino group or two neighbouring substituents a benzo-annellated radical possibly substituted by halogen,

X a sulphur atom or an imino group possibly substituted by a lower alkyl, an aryl, an aralkyl or an acyl radical,

A an amino acid or a peptide residue,

B a nitrogen protective group conventional in peptide chemistry or derived therefrom,

characterised in that, in per se known manner, one reacts compounds of the general formula II

in which

R₁, R₂, R₃, R₄ and X have the above-given meaning, with amino acids and peptides of the general formula III

in which

A and B have the above-given meaning,

or with reactive derivatives thereof.